Dioxin-like substances exhibit their toxicity by binding to the aryl hydrocarbon receptor (hereinafter referred to as “Ah receptor”) in a cell (see Ann. Rev. Pharmacol. Toxicol., 22, 517-554 (1982), and Ann. Rev. Pharmacol. Toxicol., 26, 371-399) in the following manner. If dioxin-like substances bind to the Ah receptor, this receptor is activated and translocates into the nucleus. After that, the receptor hetero-dimerizes with an Ah receptor nuclear translocator (hereinafter referred to as an “Arnt”; see Science, 252, 954-958 (1991)). This dioxin-like-substance-binding Ahr/Arnt complex binds to specific promoter element termed dioxin responsive element (DRE) (or xenobiotic responsive element (XRE); see J. Biol. Chem. 263, 17221-17224 (1988)) on a chromosome to activate the transcription of genes located downstream of the responsive element.
One method is disclosed in Fund. Appl. Toxicol., 30, 194-203 (1996). This method is the method for detecting the Ah receptor activation potency of the Lest substance, in which a reporter construct is made by linking a reporter gene, an indicator of Ah receptor activation potency, to the downstream of the responsive element, then the construct obtained thereby is introduced into a cell, and the cell is exposed to a test substance, followed by culturing, to measure the amount of reporter gene expression, thereby detecting the Ah receptor activation potency of the test substance.
Further, the activation of an Ah receptor results in increase in the expression of tyrosine hydroxylase (TH), which is the rate-limiting enzyme of dopamine biosynthesis (see Akahoshi E et al., Environ Health 7; 5; 24 (2006)). Jpn. Pat. Appln. KOKAI Publication No. 2007-202555 discloses a method for measuring the activity of a test substance against the regulatory region of a transcriptional factor having substrate binding ability. This is a method using a cell into which a vector is introduced, wherein a vector contains an enhancer sequence and a promoter sequence inserted into the upstream of luciferase genes, the enhancer sequence including 67 bp responsive sequence of a Ah receptor which mediates induction of the dioxin-responsive gene expression by dioxins and derived from mouse TH genes and the promoter sequence derived from TH genes. The cell used in this method is the one into which vectors are transfected transiently.
In view of this situation, it is desired to develop a method which is reduced in measuring time and enables a wide range of detective sensitivity stably.